A key challenge for the treatment of brain diseases is overcoming the difficulty of delivering diagnostic or therapeutic agents to specific regions of the brain. Chief amongst these obstacles is the neuroprotective Blood-Brain Barrier (BBB), both a physical and metabolic barrier to most macromolecules and small compounds. Improving our understanding of the basic molecular and cellular mechanisms that regulate brain microvasculature permeability could lead to innovative new strategies for drug or gene targeting to the injured or diseased brain. The zebrafish, a small, genetically tractable, teleost fish, possesses a BBB that shares the cardinal features of its mammalian counterpart. In this application, we will test the hypothesis that the zebrafish constitutes a novel in vivo model for BBB developmental and physiological studies in both health and disease conditions. In the first aim, in order to uncover the signals that instruct BBB formation and maintenance, we will take advantage of novel in vivo BBB labeling techniques to profile microvascular endothelial cell gene expression during both development and in adults, under physiological conditions and under chemically-mimicked hypoxia. The function of the most promising candidate genes and pathways will be analyzed via knock down approaches. In the second aim, using a herein described vascular permeability reporter transgenic line, we will perform the first high- throughput chemical screen centered on signal transduction pathways that modulate barrier permeability in the neurovascular unit. These studies have the potential to provide a better understanding of the molecular composition of the BBB endothelial cells and should pave the way towards more rationale drug delivery strategies as well as shed new lights onto how vascular permeability could be manipulated therapeutically. Taken together, they aim to provide new information as prioritized by the recently formed International Brain Barriers Society (IBBS) for the advancement of the field of BBB research and the improvement of brain disorder treatments. PUBLIC HEALTH RELEVANCE: The importance of the Blood-Brain Barrier (BBB) is illustrated by the large number of pathologies of the central nervous system (CNS) that are associated with a defective BBB. Many of the symptoms of stroke, edema, brain traumas and multiple sclerosis are due to a breakdown of the BBB that accompanies the primary insult. Furthermore, the BBB provides a stubborn obstacle in the treatment of many neurological diseases as it greatly impedes the delivery of drugs to the CNS. In this proposal we take advantage of the similarity of the zebrafish BBB to its mammalian counterpart to provide an unprecedented framework for translational research in brain barrier science.